Biomedical subjects

M Costa

Publications and source records attributed to M Costa.

At least 289 records · Page 16Linked to original sources

PubMed1993

Molecular targets of nickel and chromium in human and experimental systems.

Water-insoluble nickel compounds tend to be carcinogenic because they enter cells through phagocytosis. Highly water-soluble nickel salts, such as nickel chloride, do not enter cells well and thus are not carcinogenic in vivo. Carcinogenic nickel compounds produce selective damage to heterochromatic regions in Chinese hamster chromosomes, probably because of the high concentration of proteins and amino acids in this highly condensed genetically inactive chromatin region. Bivalent nickel ions are highly affinitive to amino acids as compared with deoxyribonucleic acid (DNA). The interaction with proteins is believed to cause this effect. In addition, when nickel binds to proteins, it can be oxidized, and DNA protein damage ensues. A new method detects DNA-protein cross-links induced by various agents, such as nickel, chromium and cis-platinum. It has demonstrated an increase in DNA-protein cross-links in cultured cells from animals treated with chromate and welders exposed to welding fumes.

Animals↗

PubMed1993

Myofibrillar breakdown and cytoskeletal alterations in heart muscle cells during invasion by Trypanosoma cruzi: immunological and ultrastructural study.

The cytoskeletal organization of normal and Trypanosoma cruzi-infected mouse embryo heart muscle cells (HMC) in primary culture was investigated using immunofluorescence and transmission electron microscopy. Fluorescent probes revealed that in the early stages of infection, up to 24 h, the HMC cytoskeleton appeared to accommodate the intracellular parasites perinuclearly, with only a few cells displaying a slight disturbance in the distribution of filaments. However, as the infection progressed (48 to 72 h), microtubules and desmin filaments were disrupted. Breakdown of myofibrils occurred in regions where the parasites were present, followed by formation of actin polygons. Using Triton X-100 treated whole cell mount, we obtained a striking preservation of the three-dimensional architecture of the cytoskeleton. Combining electron spectroscopic imaging (ESI) with contrast tuning, we detected a highly interconnected cytoskeletal network in normal cells, and a loose network in infected cells. Bundles of filaments running under and over the parasites were also observed. Our results demonstrate that T. cruzi infection induces myofibrillar breakdown and destruction of several cytoskeleton filaments in heart muscle cells.

Animals↗

PubMed1993

The inhibition of bovine serum amine oxidase by cysteamine.

Bovine serum amine oxidase (BSAO) is inhibited by cysteamine, while cystamine is a good substrate. The activity of BSAO is not restored following removal of excess cysteamine by N-ethylmaleimide, by iodoacetate or by filtration through a G-25 Sephadex column.

Animals↗

PubMed1992

Projections and chemical coding of neurons with immunoreactivity for nitric oxide synthase in the guinea-pig small intestine.

The distribution of nitric oxide synthase (NOS) immunoreactivity was investigated in the guinea-pig small intestine. There were many immunoreactive nerve cell bodies in the myenteric plexus but very few in submucous ganglia. NOS immunoreactivity was not found in non-neuronal cells except for rare mucosal endocrine cells. Abundant immunoreactive nerve fibres in both myenteric and submucous ganglia, and in the circular muscle, arose from myenteric nerve cells whose axons projected anally along the intestine. NOS immunoreactivity coexisted with VIP-immunoreactivity, but not with substance P immunoreactivity. We conclude that nitric oxide synthase is located in a sub-population of enteric neurons, amongst which are inhibitory motor neurons that supply the circular muscle layer.

Amino Acid Oxidoreductases↗

PubMed1992

Characterization of the toxicity of distamycin derivatives on cancer cell lines and rat heart.

The cytotoxicity and cardiotoxicity of benzoyl mustard (FCE 24517) and epoxamido (FCE 24561) synthetic derivatives of distamycin A were reported in the present study. The 50% inhibiting concentration (IC50) of colony formation of FCE 24517 on human SNB-19 glioblastoma, A2780 ovarian cancer and DU 145 prostate cancer was at least three times lower than that of FCE 24561; on the same cell lines the IC50 of DXR was up to 14 and 240 times higher than that of FCE 24561 and FCE 24517, respectively. Isolated rat hearts perfused with concentrations of both derivatives equivalent to their respective IC50 values did not show any significant change in ECG parameters, contractility and coronary flow. Compared to control hearts, FCE 24517 10(-6) M induced a significant increase in PR interval, reduction in + dF/dtmax, heart rate and coronary flow, while FCE 24561 10(-6) M produced a modest but significant increase in S alpha T segment and decrease in + dF/dtmax. Rats treated with FCE 24561 3, 6 or 12 mg/kg, intravenously (i.v.), once weekly for 3 weeks had a modest increase in S alpha T segment and QRS complex duration, while a slight alteration of S alpha T segment and QRS complex duration were observed in rats given FCE 24517 1 or 2 mg/kg i.v. once weekly for 3 weeks. No cardiac histologic alterations were found in hearts from rats receiving FCE 24517 or FCE 24561. For comparison, the cardiotoxicity of doxorubicin (DXR) was evaluated in the same experimental models; perfusion of hearts with DXR 10(-6) M induced severe alterations in all parameters of the isolated hearts; the administration of DXR 3 mg/kg i.v. once a week for 3 weeks was associated with a widening of the S alpha T segment and QRS complex and cardiac histologic picture was markedly altered. In conclusion, distamycin A derivatives display elevated cytotoxicity while no substantial cardiotoxicity was observed.

Animals↗

PubMed1992

Sites of action of morphine on the ascending excitatory reflex in the guinea-pig small intestine.

The effect of morphine on the ascending excitatory reflex of the circular muscle elicited by radial distension of the gut wall was studied in the isolated guinea-pig small intestine. A three compartment bath, in which an intermediate compartment divided the site of intraluminal stimulation (caudal compartment) from the site of reflex contraction recording (oral compartment), was used. Morphine (0.01-10 microM) applied independently to each compartment, caused a concentration-dependent depression (up to 90%) of the amplitude of distension-evoked reflex contractions. Concentration-response curves to morphine were shifted to the right by naloxone (30 nM) with an apparent pA2 value of about 8.5, which suggests an interaction with opioid mu-receptor subtypes. Our results indicate that morphine not only depressed transmission from excitatory motor neurons to the circular muscle but also neuro-neuronal transmission along the ascending excitatory reflex pathway.

Animals↗

PubMed1992

Selective distribution of the 66-kDa neuronal intermediate filament protein in the sensory and autonomic nervous system of the guinea-pig.

The immunohistochemical distribution of a recently identified 66-kDa neurofilament protein (NF-66) was investigated in peripheral and autonomic ganglia of the guinea-pig where it has been previously established that other neuronal intermediate filament proteins have a selective distribution. NF-66 immunoreactivity was observed in distinct subpopulations of neurons and did not coexist completely with either the neurofilament triplet or a 57-kDa intermediate filament protein (peripherin). NF-66 labelling was identical to that observed with an antibody to a 150-kDa intermediate filament or associated protein (CH1). These results further demonstrate that different neuronal intermediate filament proteins are present in selective subpopulations of neurons and that these proteins are, therefore, likely to have cell type-specific roles.

Animals↗

PubMed1992

A neurofilament protein antibody selectively labels a large ganglion cell type in the human retina.

An antibody (SMI-32) raised against the non-phosphorylated form of the neurofilament protein triplet (NFP) revealed immunoreactivity in the soma and dendritic arborization of a group of large ganglion cells in the human retina. In addition, a population of smaller somas was also faintly labeled with this antibody in the ganglion cell layer. The completely stained cells amounted to 44,000 and were non-uniformly distributed across the retina with a peak density of 100 cells/mm2 in the retinal periphery. The soma sizes increased about two-fold and dendritic field sizes about 3-fold with retinal eccentricity. The immunoreactive dendrites branched in the vitread sublamina of the inner plexiform layer. The dendritic branching pattern of these cells indicated that they correspond to the previously described shrub cells. Antibodies against NFP and neuropeptide Y showed colocalization of these markers in all of the completely stained cells.

Adult↗

PubMed1992

Ultrastructural localization of nitric oxide synthase immunoreactivity in guinea-pig enteric neurons.

Electron microscopic immunocytochemistry was used to localize immunoreactivity for nitric oxide synthase (NOS) in whole-mount preparations of myenteric plexus and circular muscle from guinea-pig ileum. NOS immunoreactivity was patchily distributed in myenteric neurons and was not specifically associated with any subcellular organelle or with the plasma membrane. This localization leaves unanswered the question of how nitric oxide is stored and released. NOS immunoreactive fibres in the circular muscle were found closer than 100 nm to muscle cells. NOS immunoreactive nerve fibres made synaptic contacts with NOS immunoreactive and non-immunoreactive enteric neurons. These results indicate that nitric oxide may regulate the activity of both myenteric neurons and smooth muscle.

Amino Acid Oxidoreductases↗

PubMed1992

Projections and pathways of submucous neurons to the mucosa of the guinea-pig small intestine.

Double-labelling immunohistochemistry and retrograde transport of the carbocyanine dye, DiI, were used to establish the pathways of submucous neurons to the mucosa of the guinea-pig small intestine. Following the application of DiI to a villus, DiI-labelled nerve cell bodies were found in the submucous plexus up to 8.3 mm circumferentially and 3.8 mm longitudinally. The size of each of the four characterised classes of submucous neurons was determined and their distributions and projections mapped. Cells characterised by vasoactive intestinal polypeptide immunoreactivity accounted for 52% of DiI-labelled cells and had the longest projections. Cells characterised by neuropeptide Y (19%) or by calretinin immunoreactivity (13% of all DiI-labelled neurons) had relatively short projections and cells with substance P immunoreactivity (20%) had intermediate lengths of projection. When DiI was applied directly to the submucous plexus, filled neurons of all classes had significantly shorter projections, indicating that they must run for considerable distances in other pathways to the mucosa, probably via the non-ganglionated plexus. On average, each villus is innervated by at least 70 submucous neurons. From quantitative estimates there are 9 submucous neurons per villus. Thus, each submucous neuron is likely to supply about 8 villi. This demonstrates a high degree of convergence and divergence in the innervation of the mucosa.

Animals↗

PubMed1992

Reduced cardiotoxicity and increased cytotoxicity in a novel anthracycline analogue, 4'-amino-3'-hydroxy-doxorubicin.

The acute and chronic cardiotoxicity and cytotoxicity of the novel doxorubicin (DXR) derivative 4'-amino-3'-hydroxy-DXR were compared with those of 4'-deoxy-DXR and DXR. In the acute cardiotoxicity study, the ECG and hemodynamic changes recorded in anesthetized rats that had been treated i.v. with 10 mg/kg 4'-amino-3'-hydroxy-DXR or 8.6 mg/kg 4'-deoxy-DXR were significantly less severe than those caused by 13 mg/kg DXR. In the chronic cardiotoxicity study, rats received 3 weekly i.v. injections of 3 mg/kg DXR, 3 mg/kg 4'-amino-3'-hydroxy-DXR, or 2 mg/kg 4'-deoxy-DXR during the first 14 days of the study and were observed for an additional 35-day period. DXR induced severe cardiomyopathy that was characterized by ECG changes in vivo (S alpha T-segment widening and T-wave flattening) and by impairment of the contractile responses (Fmax, +/- dF/dtmax) to adrenaline of hearts isolated from treated animals. 4'-Deoxy-DXR caused a progressive enlargement of the S alpha T segment in vivo and a significant impairment of the -dF/dtmax value in vitro, which were less severe than those produced by DXR. The least cardiotoxic drug was 4'-amino-3'-hydroxy-DXR, which induced minor ECG changes without causing significant alterations in the contractile responses of isolated hearts to adrenaline. On the basis of the drug concentration required to inhibit 50% of the colony formation (IC50) of cell lines in vitro, 4'-amino-3'-hydroxy-DXR was less active than 4'-deoxy-DXR but at least twice as active as DXR against human cancer and murine transformed cell lines. These data indicate that 4'-amino-3'-hydroxy-DXR is significantly less cardiotoxic and more cytotoxic than DXR.

Animals↗

PubMed1992

Reducing doxorubicin cardiotoxicity in the rat using deferred treatment with ADR-529.

The purpose of this study was to evaluate the optimal timing of ADR-529 administration to protect rats treated with doxorubicin (DXR) against drug-induced cardiotoxicity. Complete electrocardiographic monitoring (QRS complex, S alpha T segment and T wave) and the histopathological analysis of cardiac tissue were used to assess the degree of heart damage produced in female rats treated with ten i.v. doses of 1 mg/kg DXR over a period of 15 weeks; body-weight increase and survival were also analyzed to evaluate the toxicity of treatments. Cardiac alterations induced by DXR were compared with those occurring in animals receiving 20 mg/kg i.v. ADR-529 at 30 min prior to DXR administration, starting at the first, third, or sixth DXR dose and given until the end of the study (15th week). Rats treated with DXR were severely cardiomyopathic, showing progressive and irreversible ECG alterations (QRS-complex and S alpha T-segment widening and T-wave flattening) and marked degeneration of the myocardium (myocyte vacuolation, myofibrillar loss, and endomyocardial fibrosis). The most effective cardiac protection was provided by the administration of ADR-529 beginning with the first or third DXR dose. Delaying treatment with ADR-529 until the sixth DXR dose resulted in a significant reduction in its therapeutic action on heart damage. A significant difference in body-weight increase and survival was observed between the treatment groups: ADR-529 injected prior to the first DXR dose significantly protected animals from DXR toxicity, but this schedule was significantly more toxic than the administration of ADR-529 beginning with the third or sixth DXR dose. Taking into account the degree of cardiac protection and the toxicity of combination treatments, the results of the present study demonstrate the superiority of ADR-529 given prior to the third DXR dose over the other schedules tested. This finding suggests that significant protection against DXR-induced chronic cardiotoxicity in the rat can be obtained using deferred treatment with ADR-529.

Animals↗

PubMed1992

Identification of motor neurons to the longitudinal muscle of the guinea pig ileum.

Motor neurons that innervate the longitudinal muscle of the guinea pig ileum were identified by retrograde transport from the longitudinal muscle plexus in organotypic culture. Motor neurons had short projections, less than 3.5 mm long, and never had Dogiel type II morphology; most labeled neurons had morphological characteristics of Dogiel type I neurons. Immunoreactivity for choline acetyltransferase was present in 97% of retrogradely labeled nerve cell bodies, reflecting the dominant cholinergic input to the longitudinal muscle layer. Substance P immunoreactivity was present in 48% of motor neurons, indicating that it or a similar tachykinin that mediates noncholinergic excitatory transmission is likely to be released by a subset of cholinergic motor neurons. This strongly suggests that the difference in frequency dependence of substance P and acetylcholine release is attributable to different release mechanisms rather than to activation of separate populations of motor neurons. Immunoreactivity for the calcium-binding protein calretinin was present in 87% of longitudinal muscle motor neurons. The neurochemical coding of longitudinal muscle motor neurons indicated that they constitute about one quarter of all myenteric neurons and are distinct from circular muscle motor neurons.

Animals↗

PubMed1992

Forward mutations and DNA-protein crosslinks induced by ammonium metavanadate in cultured mammalian cells.

Ammonium metavanadate yielded a dose-dependent increase in mutation frequency at the V79 hprt locus following a 24-h exposure period in serum-free F12 medium. Vanadate also increased the mutation frequency of V79 cells by exposure of cells in salts-glucose medium, but these effects were not as striking, or as dose-dependent as they were in serum-free F12 medium. Ammonium metavanadate enhanced the mutation frequency in a V79 variant containing a transfected bacterial gpt gene. These cells are known to be more responsive to oxidative type mutations, and to mutations involving deletions. Although the absolute level of mutations was greater in these cells with ammonium metavanadate, so was the background, and these cells did not exhibit an enhanced mutagenic response to vanadate when compared to the wild-type V79 cells. The vanadate results were compared to a positive control potassium chromate, which exhibited a dose-dependent increase in mutation frequency. Ammonium metavanadate induced DNA-protein crosslinks formation in both Chinese hamster ovary and human MOLT4 cells, and the role of these relatively unrepaired genetic lesions in the mutations produced by vanadate and chromate are discussed.

Animals↗

PubMed1992

Potentiation of sodium chromate(VI)-induced chromosomal aberrations and mutation by vitamin B2 in Chinese hamster V79 cells.

The effect of vitamin B2, which is capable of reducing chromium(VI) to chromium(V), on chromosomal aberrations and mutation caused by Na2CrO4 was investigated in Chinese hamster V79 cells. Pretreatment with 200 microM vitamin B2 (riboflavin) for 24 h prior to exposure to Na2CrO4 (2.5-5 microM) resulted in an increase of metal-induced chromosomal aberrations and mutation at the HGPRT locus. These and other previous studies suggest that vitamin B2 enhances the clastogenic and mutagenic action of chromate compounds, through its ability to directly reduce chromium(VI) in cells.

Animals↗

PubMed1992

The neurofilament triplet is present in distinct subpopulations of neurons in the central nervous system of the guinea-pig.

It is commonly assumed that most, if not all, neurons contain the intermediate filament protein class known as the neurofilament protein-triplet. The following study investigated the distribution of neurofilament protein-triplet immunoreactivity in selected regions of the guinea-pig central nervous system using monoclonal antibodies directed against phosphorylation-independent epitopes on the three subunits under optimal tissue processing conditions. Neurofilament protein-triplet immunoreactivity was present in distinct subpopulations of neurons in the cerebellar cortex, neocortex, hippocampal formation, retina, striatum and medulla oblongata. In many of these regions, labelled neurons represented only a small proportion of the total. The selective distribution of this intermediate filament protein class was confirmed in double-labelling experiments using antibodies to the neurofilament protein-triplet in combination with antibodies to other neuronal markers. The distribution of neurofilament protein-triplet immunoreactivity also correlated with the distribution of staining observed with a silver impregnation method based on Bielschowsky. The present results in combination with previous observations have demonstrated that the neurofilament protein-triplet is found in specific subclasses of neurons in different regions of the nervous system. Content of this intermediate filament protein class does not appear to be correlated with neuronal size or length of projection. These results also suggest that the selectivity of staining between neuronal classes observed with classical silver impregnation methods may be due to the presence or absence of the neurofilament protein-triplet. The present results may also provide a new perspective on the basis of the selective vulnerability of neurons in degenerative diseases.

Animals↗

PubMed1992

Analysis of residual amino acid--DNA crosslinks induced in intact cells by nickel and chromium compounds.

Chinese hamster ovary cells were incubated with radioactive amino acids, the DNA was isolated by standard proteinase K/phenol/chloroform extraction and residual amino acids complexed to the DNA were examined as an index of metal induced DNA-protein crosslinks. Using this method, both chromate and nickel caused residual histidine and cysteine to be complexed with the DNA isolated from metal-treated cells. In the case of chromate, a number of amino acids were studied and Tyr, Thr and Cys were found to be complexed to DNA at a level (above the untreated control) that was statistically significant. Stability studies indicated that some of the chromate-induced DNA-protein complexes were mediated by direct participation of chromium(III), whereas others that were resistant to dissociation by EDTA and mercaptoethanol did not seem to involve direct chromium(III) participation. A significant portion of the cysteine complexed to DNA by chromate was believed to involve glutathione since treatment of cells with cycloheximide did not decrease chromate-induced cysteine-DNA crosslinks. In the case of nickel, most of the stable DNA-protein crosslinks did not involve direct metal participation and were probably oxidatively mediated by Ni(II)/Ni(III) redox cycling. These findings present new methodology for analysis of DNA-protein crosslinks by examination of residual amino acids associated with the DNA. This method should be highly sensitive and will yield important information about the mechanism of metal-induced DNA-protein crosslinks.

Amino Acids↗

PubMed1992

Analysis of the binding sites of chromium to DNA and protein in vitro and in intact cells.

Previous studies have examined Cr(III), or CrO4 reduced to Cr(III), binding in vitro to DNA. However, there have been few studies examining chromate binding to DNA in intact cells. Treatment of intact cells with chromate (Na2(51)CrO4) resulted in chromium (Cr) binding to DNA. The binding of Cr to DNA was much more stable when more residual peptide/amino acids were associated with DNA. A substantial portion of the Cr bound to DNA was released by treatment with EDTA, suggesting that trivalent Cr was the major oxidation state of Cr bound to DNA. Cr(III) stimulated the formation of amino acid-DNA and protein-DNA complexes in vitro. Tyrosine and cysteine exhibited the highest activity in being complexed to DNA by Cr(III) in vitro, while histidine, methionine and threonine also exhibited more activity than any other amino acid. Similar results were found in intact cells. The activity of proteins complexed to DNA by trivalent Cr depended upon the content of these reactive amino acids. Thus, bovine serum albumin was more active than actin, which in turn was more active than histones. These and other studies presented suggested that Cr(III) was involved directly in the formation of DNA-protein complexes in intact cells, unlike other metals such as Ni(II), which are thought to form DNA-protein cross-links catalytically and not participate directly in the complex. The majority of trivalent Cr associated with DNA was bound to the phosphate backbone without exhibiting any base specificity. Collectively, these results indicate that trivalent Cr creates DNA protein crosslinks by binding with reactive amino acids (i.e. cysteine, tyrosine or histidine) and linking these to the phosphate backbone of DNA.

Amino Acids↗